Understanding the mechanisms that drive and organize protein adsorption onto biological membranes is fundamental to clarifying membrane-mediated protein function. It is known that cytochrome c is a small, membrane-associated protein playing a critical role in the energy production machinery in the mitochondria. Recently, it was found that cytochrome c release from the inner mitochondria membrane, where it primarily resides, is the first step in the apoptotic pathway. The inner mitochondria membrane is rich in anionic lipids but the exact functional localization of the protein remains obscure despite the number of past efforts to resolve the question using indirect methodologies. Our staff, in collaboration with Dr. Allen Minton of NIDDK, examined the adsorption of the protein to anionic supported bilayers under low ionic conditions. It appears that the protein does not adsorb to the bilayer surface yet its interaction with the bilayer results in a significant increase of bilayer fluidity. This result has two important implications: 1. Cytochrome c in low ionic strength conditions probably inserts into the hydrophobic core of anionic lipid bilayers resulting in a significant perturbation of the bilayer structure (hence its increased fluidity). 2. The AFM can be used, in the force mode, to probe and quantify fundamental thermodynamic properties of bilayers (yield strength, line tension, elasticity modulus, etc.). We are examining both implications further by continuing the study of cytochrome c adsorption to bilayers that closer resemble the inner mitochondria membrane as well as bilayers under varying ionic conditions; by complementing the AFM with the Raman spectroscopy technique that will ascertain the presence or absence of the protein and will give indications of conformational/structural changes associated with the interaction; and, by further developing the AFM technique for the measurement of fundamental thermodynamic parameters of bilayers in general.